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Sample pretreatment method of endogenous brassinosteroids in plant sample

A brassinosteroid and sample pretreatment technology, applied in the field of analytical chemistry, can solve the problems of burden, high solvent consumption and high cost, and achieve the effects of high selectivity, simple method and easy operation.

Active Publication Date: 2013-07-17
武汉绿剑可瑞信科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Most of these methods require dozens of grams of fresh plant tissue. Such a large sample volume is difficult to achieve for some precious samples, and it also brings problems such as large solvent consumption and high cost in the pretreatment process.
In addition, the above pretreatment method only achieves the removal of impurities through the difference in hydrophobicity between BR and impurities, which will inevitably cause co-elution of impurities and BRs, which will inevitably bring a burden to the subsequent liquid phase separation, thereby reducing the mass spectrometric ionization of BRs. efficiency

Method used

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  • Sample pretreatment method of endogenous brassinosteroids in plant sample
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  • Sample pretreatment method of endogenous brassinosteroids in plant sample

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Accurately weigh 1g of plant tissue, grind it into powder with liquid nitrogen, transfer it to a 10mL centrifuge tube, add isotope internal standard [ 2 h 3 ] brassinolide and [ 2 h 3 ] Brassinosterone 2ng each, after standing for a while, add 5mL acetonitrile and mix well. Place in a -18°C refrigerator for extraction for 12 hours. Take the supernatant by centrifugation, add 0.5 g of sodium chloride to the supernatant to promote the separation of the solution, and discard the aqueous phase. Then add 0.5 g of anhydrous magnesium sulfate to remove the remaining moisture. Centrifuge to remove the supernatant and discard the insoluble matter. The supernatant was extracted by pre-activated graphitized carbon black / ethylenediamine-N-propyl bonded silica double-layer solid-phase extraction, desorbed with methanol / acetonitrile (1 / 1, v / v, 2mL); collected and loaded The effluent and desorbent were blown dry. Dissolve the dried sample residue with a buffer solution (phospha...

Embodiment 2

[0027] Accurately weigh 1g of plant tissue, grind it into powder with liquid nitrogen, transfer it to a 10mL centrifuge tube, add isotope internal standard [ 2 h 3 ] brassinolide and [ 2 h 3 ] Brassinosterone 2ng each, after standing for a while, add 5mL acetonitrile and mix well. Place in a -18°C refrigerator for extraction for 12 hours. Take the supernatant by centrifugation, dry the supernatant with nitrogen, and dissolve the residue with acetonitrile. The dissolved sample was passed through a pre-activated carbon nanotube / pure silica gel double-layer solid-phase extraction column, desorbed with methanol (2 mL); the sample effluent and desorbed liquid were collected and dried. Dissolve the dried sample residue with buffer solution (phosphate buffer solution, 20mM, pH11.0, 1mL), pass the solution through a pre-activated commercial phenylboronic acid-bonded silica gel solid-phase extraction column, and use buffer solution (phosphate buffer solution, 20mM, pH9.0, 1mL) to ...

Embodiment 3

[0029] Accurately weigh 1g of plant tissue, grind it into powder with liquid nitrogen, transfer it to a 10mL centrifuge tube, add isotope internal standard [ 2 h 3 ] brassinolide and [ 2 h 3] Brassinosterone each 1ng, after standing for a while, add 5mL acetonitrile and mix well. Place in a -18°C refrigerator for extraction for 12 hours. Take the supernatant by centrifugation, add 0.5 g of sodium chloride to the supernatant to promote the separation of the solution, and discard the aqueous phase. Then add 0.5 g of anhydrous magnesium sulfate to remove the remaining moisture. Centrifuge to remove the supernatant and discard the insoluble matter. Add the mixed material of graphitized carbon black, C18 bonded silica gel and magnetic particles to the supernatant, vortex and mix for 5 minutes, and collect the loading effluent. The brassinosterol adsorbed on the material was then desorbed with 2 mL of methanol. Combine the sample effluent and desorbed solution, and evaporate ...

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Abstract

The invention discloses a sample pretreatment method of the endogenous brassinosteroids in a plant sample. The method comprises the following steps that: firstly, the endogenous brassinosteroids are extracted by a solvent; and then the impurities of the solution obtained in the previous step are further removed with boron-affinity material as an extraction medium, so that the sample pretreatment of the endogenous brassinosteroids is realized. The method has the advantages of simplicity, rapidness, solvent conservation and simplicity in operation. Besides, according to the method, high selectivity is shown in the aspect of removing endogenous impurities of plant extracting solution and enrichment and purification of low-content endogenous brassinosteroids can be realized.

Description

technical field [0001] The invention relates to a sample pretreatment method for endogenous brassinosterol in plant samples, belonging to the field of analytical chemistry. Background technique [0002] Brassinosteroids (BRs) are a class of polyhydroxylated sterol compounds, which are found in extremely low levels in plants, but they regulate a series of physiological and metabolic processes in plants. In recent years, studies on the physiological functions of brassinosterols, such as the biosynthesis, degradation and metabolic pathways of BRs, have received extensive attention. However, due to the complex matrix and extremely low brassinosterol content in plant extracts, the development of these studies is limited. Therefore, establishing an efficient sample pretreatment method combined with a sensitive detection method is the key to the detection of endogenous BRs. [0003] The reported pretreatment methods of brassinosteroids in plants include liquid-liquid extraction, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N1/28G01N1/34
Inventor 冯钰锜丁俊毛利菁
Owner 武汉绿剑可瑞信科技有限公司
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